The relatively poor stability to aqueous eluents at elevated pH of chromatography column packing materials containing organic coatings covalently attached directly to the silanol groups on the surface of silicon oxide (silica) is well known. Chromatography columns prepared with these packings generally begin to lose resolving power when these covalent bonds are hydrolyzed from the packing surfaces to a significant degree. This degeneration results in reduced chromatographic column utility manifested by reduced periods of use or diminished separation capability. There is a need for chromatographic packings which are relatively inexpensive, each to manufacture, and maintain acceptable functional properties over extended periods to make long-term, repeated uses possible.
A method for improving the water-durability of controlled pore glass (CPG)-immobilized enzymes was introduced in U.S. Pat. No. 3,783,101, issued Jan. 1, 1974 to Tomb et al. Inorganic carriers, for example, CPG were coated with various metal oxides, including zirconium oxide, and fired at between 200.degree. C. and 800.degree. C. to form a continuous metal oxide layer on the CPG surface. The metal oxide layer was silanized for subsequent functional group immobilization. A continuous barrier was formed between the CPG and the silane coupling agent by the metal oxide layer so that no interaction of glass and silane occured. Enzymes immobilized on the silanized, metal oxide coated CPG exhibited longer half-lives of activity than enzymes immobilized on either uncoated CPG or silanized, uncoated CPG. This improvement is said to be caused by the continuous metal oxide coating over the inorganic carrier surface which inorganic carrier was less water durable than the metal oxide.
U.S. Pat. No. 3,910,851, issued Oct. 7, 1975 to Messing, discloses that pore size in CPG can be optimized with respect to molecular sizes of substrate added to, and/or enzymes immobilized on, CPG, to provide an improved surface area for enzyme immobilization without risking excessive enzyme losses from pore surfaces due to flow-through turbulence. Uniform pore size porous inorganic bodies were prepared from sol particles of SiO.sub.2, Al.sub.2 O.sub.3 or TiO.sub.2 using water-soluble zirconium compounds as binder. A slurry of sol particles having a specified diameter range was treated with a zirconium binder, the mixture dried and fired at temperatures in the range 450.degree. to 800.degree. C. When SiO.sub.2 sol particles were used, for example, the firing step provided zirconium silicates to impart physical stability. While the use of zirconium binders imparts control of resulting pore size distributions when slurries are dried and fired below sintering temperatures, the process results in solid aggregates unsuitable for chromatography.
It has been found that the stabilized bodies, especially the stabilized microspheres produced by this invention, comprising porous silica microspheres with surfaces partially covered by metal oxides, retain the highly controlled pore size distribution of the starting material silica microspheres. The discontinuous metal oxide coating imparts hydrolytic stability to subsequently applied organosilane coatings over the pH range 3.0 to 9.0. The organo-silane coatings can be modified by known methods to produce a wide variety of stable chromatographic packings.